(i) Field of the Invention:
The present invention relates to maize starches with an apparent amylose content between 35 wt. % and 90 wt. % and with improved processing properties relative to conventional high-amylose maize starches. Furthermore, the present invention relates to maize meals and foodstuffs containing these maize starches or maize meals. In addition the present invention relates to methods of production of said maize starches/maize meals and maize plants which synthesize these maize starches. Moreover, the present invention relates to wheat starches with an apparent amylose content between 35 wt. % and 90 wt. % and improved processing properties, and wheat meals and foodstuffs containing these wheat starches or wheat meals. In addition, the present invention relates to methods of production of said wheat starches/wheat meals and wheat plants that synthesize these wheat starches.
(ii) Description of the Related Art:
Apart from oils, fats and proteins, polysaccharides are the main renewable raw materials from plants. Along with cellulose, starch occupies a key position among the polysaccharides, and is one of the most important storage substances in higher plants.
Furthermore, from the standpoint of nutritional physiology, starch is an important constituent of the human and animal diet. The structural characteristics of the starch present in foodstuffs can influence the functional (e.g. water binding capacity, swelling power), nutritional-physiological (e.g. digestibility, influence of the foodstuff on the glycemic index) or structure-forming (e.g. resistance to cutting, texture, stickiness, processability) properties of the most varied foodstuffs. Foodstuff compositions therefore often contain a starch with specified structural characteristics, which determine the desired properties of the foodstuff in question. The properties of foodstuffs containing starch-storing plant tissues (e.g. cereals, fruits, meals), can also be influenced by the starch contained in the plant tissues.
The polysaccharide starch is a polymer of chemically uniform basic building blocks, the glucose molecules. However, it is a very complex mixture of various molecular forms, which vary with respect to their degree of polymerization, the presence of branchings of the glucose chains and their chain lengths, and furthermore, may be modified, e.g. phosphorylated. Therefore starch is not a uniform raw material. In particular we distinguish amylose, an essentially unbranched polymer of alpha-1,4-glycosidically linked glucose molecules, from amylopectin, which is a complex mixture of variously branched glucose chains. The branchings arise because of the presence of additional alpha-1,6-glycosidic linkages. In typical plants used for industrial starch production or as foodstuffs, such as maize, rice, wheat or potato, the starch that is synthesized consists of approx. 20%-25% amylose and approx. 70%-75% amylopectin.
Various methods are available for determining amylose content, and they can lead to different numerical values of the amylose content for one and the same starch. Many of these methods are based on the iodine-binding capacity of amylose, which can be determined potentiometrically (Banks & Greenwood, in W. Banks & C. T. Greenwood, Starch and its components (pp. 51-66), Edinburgh, Edinburgh University Press), amperometrically (Larson et al., Analytical Chemistry 25(5), (1953), 802-804) or spectrophotometrically (Morrison & Laignelet, J. Cereal Sc. 1, (1983), 9-20). The amylose content can also be determined calorimetrically by DSC (differential scanning calorimetry) measurements (Kugimiya & Donovan, Journal of Food Science 46, (1981), 765-770; Sievert & Holm, Starch/Starke 45 (4), (1993), 136-139). It is also possible to determine the amylose content using SEC (size exclusion chromatography) chromatography of native or debranched starch. This method was recommended in particular for determining the amylose content of genetically engineered starches (Gérard et al., Carbohydrate Polymers 44, (2001), 19-27).
The functional, nutritional-physiological or structure-forming properties of starch, such as solubility, retrogradation behavior, water-binding capacity, film-forming properties, viscosity, gelatinization properties, freezing/thawing stability, acid resistance, gel strength, swelling power, digestibility, starch granule size of starches are influenced by, among other things, the structural characteristics of the starch, such as the amylose/amylopectin ratio, the molecular weight of the glucose polymers, the pattern of side chain distribution, the phosphate content, the lipid content or the protein content.
Using classical breeding methods, precise alteration of the structural/functional properties of the starch synthesized in plants is very difficult and is only possible for selected structural characteristics. An alternative to breeding methods is the precise modification of starch-producing plants by the methods of genetic engineering. However, a precondition for this is the identification and characterization of the enzymes involved in starch synthesis and/or starch modification and their subsequent functional analysis in transgenic plants.
Maize mutants with an increased amylose content relative to wild-type maize plants are known and are designated as “amylose extenders” or “ae” for short. Amylose extender (ae) maize mutants were described for example in Vineyard and Bear (Maize Genet Coop Newsletter 26: 5 (1952), who describe the reference allele ae1-Ref, and in Moore and Creech (Genetics 70, (1972), 611-619), Garwood et al. (Cereal Chemistry 53(3), (1976), 355-364) and Hedman and Boyer (Biochemical Genetics 21 (11/12), (1983), 1217-1222).
Maize plants (cells) that have an “amylose extender mutation” show a mutation of the gene of the starch branching enzyme IIb from maize (abbreviation “BE IIb” or “SBE IIb”), which is also designated as amylose extender gene. This mutation leads to a decrease in SBE IIb enzyme activity in the endosperm of these maize plants compared to the BE IIb activity in the endosperm of wild-type maize plants. Preferably this mutation of BE IIb in maize plants (cells) has the effect that SBE IIb activity is no longer detectable (e.g. Fisher et al., Plant Physiol. 110, (1996), 611-619, in particular FIG. 4; Hedman and Boyer, Biochemical Genetics 21 (11/12), (1983), 1217-1222, in particular Table 1).
The “amylose extender mutation” is as a rule a recessive mutation of the amylose extender 1 locus.
Amylose extender mutants synthesize a high-amylose maize starch, which has an increased apparent amylose content compared with wild-type maize plants (plant cells), which as a rule is between 45 and 75 wt. %, depending on the genetic background, cultivation conditions and method of amylose determination.
Maize starches with the product designation Amylogel®, Hylon® V and Hylon® VII are available commercially and as a rule have an apparent amylose content of approx. 50% or 70% (Shi et al., Journal of Cereal Science 27, (1998), 289-299).
In contrast to wild-type maize starches, these high-amylose maize starches have a greatly reduced swelling power (Senti and Russell, Tappi Vol. 43 No. 4, (1960), 343-349; Shi et al., Journal of Cereal Science 27, (1998), 289-299).
Aqueous suspensions of the high-amylose maize starches show no development of viscosity in viscosity analysis (e.g. RVA analysis) (Senti and Russell, Tappi Vol. 43 No. 4, (1960), 343-349). Native high-amylose maize starches so far only undergo gelatinization in a very energy-intensive and therefore cost-intensive method in an autoclave at elevated temperature and pressure, or they require subsequent chemical modification.
Based on the aforementioned properties, the high-amylose maize starches are very difficult to process, so that the range of applications in which the advantageous properties of the native high-amylose maize starches (e.g. very good film-forming properties, good gel-forming properties, use as resistant starch with prebiotic action) can be utilized is still very limited. Therefore there is a great demand for high-amylose maize starches with improved processing properties, such as increased solubility in hot water, increased swelling power and capacity for viscosity development in aqueous suspensions under normal conditions. High-amylose wheat starches with good processing properties are also not available.